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Mol. Cell. Biol., 01 1995, 488-496, Vol 15, No. 1
Copyright © 1995, American Society for Microbiology

Lack of an effect of the efficiency of RNA 3'-end formation on the efficiency of removal of either the final or the penultimate intron in intact cells

D Nesic, J Zhang and LE Maquat
Department of Human Genetics, Roswell Park Cancer Institute, Buffalo, New York 14263.

Evidence exists from studies using intact cells that intron removal can be influenced by the reactivity of upstream and downstream splice sites and that cleavage and polyadenylation can be influenced by the reactivity of upstream splice sites. These results indicate that sequences within 3'-terminal introns can function in the removal of upstream introns as well as the formation of RNA 3' ends. Evidence from studies using intact cells for an influence of RNA 3'-end formation on intron removal is lacking. We report here that mutations within polyadenylation sequences that either decrease or increase the efficiency of RNA 3'-end formation have no effect on the efficiencies with which either the 3'-terminal or the penultimate intron is removed by splicing. Northern (RNA) blot hybridization, RNase mapping, and an assay that couples reverse transcription and PCR were used to analyze the effects of deletions and a substitution of the polyadenylation sequences within the human gene for triosephosphate isomerase (TPI). TPI pre-mRNA harbors six introns that are constitutively removed by splicing. Relative to normal levels, each of the deletions was found to reduce the nuclear and cytoplasmic levels of TPI mRNA, increase the nuclear level of unprocessed RNA 3' ends, and decrease the nuclear level of processed RNA 3' ends.(ABSTRACT TRUNCATED AT 250 WORDS)

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